3D High Throughput Model to Predict Drug Efficacy in Fibrosis Progression vs Reversal

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ABSTRACT: 3D High Throughput Model to Predict Drug Efficacy in Fibrosis Progression vs Reversal Idiopathic pulmonary fibrosis (IPF) is the most relentlessly progressive and fatal fibrotic lung disorder, which disproportionately affects the elderly. Although two drugs have recently gained FDA-approval for IPF, these drugs only moderately slow the progression of lung decline and do not improve quality of life for patients. There are no available therapies that can `reverse' fibrosis. Despite efforts by numerous groups to develop IPF treatments, progress has been aggravatingly slow. This proposal focuses on two possible reasons for these difficulties: (1) Current pre-clinical screening models fail to reliably predict the success of drug candidates in humans, and (2) Although IPF is widely regarded as an age-related disease, drug treatments have not targeted age-associated pathologic mechanisms. The existing paradigm, that pathologic fibrosis is a “fibro-proliferative” process, has not led to effective IPF treatments. This proposal integrates expertise in fibroblast aging and novel IPF therapeutics in development (Hecker lab) with cutting edge technologies for microscale bioprinting and 3D cell assays (Takayama lab) to develop a high throughput phenotypic cellular screening assay to determine efficacy for fibrosis reversal. The proposed studies will utilize normal “control”, aged “senescent”, and IPF human lung fibroblasts in small numbers to bioengineer a high-throughput phenotypic assay that will evaluate fibrosis over a 21 day period. An aqueous two phase system (ATPS) bioprinting of these cells will be used to create microscale contraction assays that are several order of magnitude smaller in volume compared to conventional assays. Importantly, the project will repeatedly micro-print fresh collagen around already contracted cell-laden gels to enable repeated contractions over 21 days. The proposed model will enable the first high-throughput phenotypic screening assay with the capability to determine a drug candidate's efficacy for fibrosis progression and reversal. The new cellular assay will be validated for its ability to identify fibrosis reversal drugs using “Noxindoline” a highly selective Nox4 inhibitor that is currently in preclinical development by the Hecker lab. Noxindoline was identified by the Hecker lab through studies of age-dependent alterations in Nox4 that results in a sustained redox imbalance, and promotes senescence and apoptosis-resistance of myofibroblasts. The proposal hypothesizes that current therapies (Nintedanib and Pirfenidone) will inhibit the progression of pro-fibrotic phenotypes (but not reversal), whereas treatment with Noxindoline will promote the reversal of established pro-fibrotic phenotypes. The aims are: Aim1: Develop high throughput bioprinted cellular assay for fibrosis progression using non-senescent cells Aim2: Monitor fibrosis progression and reversal of senescent cells and IPF patient cells